PROJECT SUMMARY The process of mRNA translation lies at the heart of cellular metabolism. In cancers, many different oncogenic signaling pathways converge on the translation machinery to increase translational flux. While our knowledge of translational regulation is steadily increasing, it is unclear how oncogenic translation supports and promotes the malignant state. The goal of this project is to investigate the signature of oncogenic translation, explore the translational vulnerabilities associated with cancer progression, and develop therapeutic strategies targeting aberrant translation of tumor cells. Using an inducible RasG12V cell model, we discovered that oncogenic translation is characterized by relaxed initiation pausing. Remarkably, mRNA methylation in the vicinity of start codons influences the behavior of initiating ribosomes, thereby causing dynamic initiation pausing. Intriguingly, oncogenic RAS signaling reduces global m6A levels, suggesting a crucial role of mRNA epigenetics in malignant transformation and tumor growth. These findings led to the central hypothesis that oncogenic translation not only involves enhanced cap-recognition, but also relies on relaxed initiation pausing to achieve the maximal translational output. To test this hypothesis, the following Aims are proposed: 1) Determine the causal relationship between mRNA methylation and initiation pausing; 2) Target oncogenic translation by modulating initiation pausing. By integrating innovative approaches into fundamental studies of RNA biology, the proposed studies will open up new avenues of research in the field of mRNA epigenetics, protein translation, and cancer biology. The mechanistic insights we gain from this study will provide paradigms for better understanding of translational control in tumor growth. Ultimately, it will set the stage for novel therapeutic strategies in cancer treatment.